A fundamental question in biology is how one cell divides to yield two progeny with different identities. Like eukaryotic cells, bacteria undergo complex life cycles and often produce daughter cells with distinct shapes and properties. In this work, Caulobacter crescentus is used as a model system for the study of asymmetry coupled to the division cycle. In this bacterium, each division produces two morphologically distinct daughter cells, a non-motile cell with a stalk that attaches to surfaces and a motile swarmer cell with a flagellum that propels it through the water. The stalked cell immediately begins a new round of chromosome replication and division, but the swarmer cell is unable to initiate DMA replication until it differentiates into a stalked cell. This complex cell division cycle is orchestrated by a network of two-component signal transduction proteins. The response regulator CtrA is a transcription factor that controls the expression of many cell cycle- regulated genes but also blocks DNA replication by binding to the origin of replication. CtrA activity is required for cell viability but it must be temporarily eliminated in stalked cells to permit the initiation of chromosome replication. CtrA activity is indirectly opposed by the essential response regulator DivK. Phosphorylated DivK results in a decrease in CtrA activity, which ultimately allows chromosome replication in the stalked cell. Thus, phosphorylation of DivK is essential for viability. DivK is known to be activated by the histidine kinase (HK) DivJ but because DivJ is dispensible, DivK must be phosphorylated by another HK or small molecule phosphodonor in Caulobacter. We propose a whole-genome approach to identify other HKs that could participate in DivK phosphorylation. Specifically, we aim to delete the gene for each of the 59 non-essential HKs in combination with a divJ deletion. This approach has yielded a likely candidate which we have tentatively named DivM other candidates must still be ruled out. We also aim to characterize the terminal phenotype of cells lacking both DivJ and DivM, determine the location and activity of DivM during the cell cycle, and elucidate the phosphorylation pathway from DivM to DivK. PUBLIC HEALTH RELEVANCE: The work proposed here is integral to achieving a complete understanding of the regulatory cascade leading to cell-cycle progression and differentiation in Caulobacter. Comprehension of the regulatory cascade could have far-reaching implications because many of the mechanisms discovered in Caulobacter are conserved among other species with important roles in agriculture, biowarfare, biosensing, and bioengineering. In addition, elucidating the basic mechanisms involved in bacterial cell cycle progression will generate key insights into prokaryotic cell biology, which will in turn help to identify new targets for antibacterial drug discovery.